Quantification and functional analysis of chemotaxis by laser scanning cytometry.
Quantification and functional analysis of chemotaxis by laser scanning cytometry.
复制标题
通过激光扫描细胞术对趋化性进行定量和功能分析。
DOI：
10.1002/cyto.a.20118
复制
发表时间：
2005
期刊：
影响因子：
--
通讯作者：
Moldovan,NicanorI
中科院分区：
文献类型：
--
作者：
Butt,OmarI;Krishnan,Padmavathy;Kulkarni,SumantS;Moldovan,Leni;Moldovan,NicanorI
背景趋化性测定的评价传统上依赖于繁琐且有时不准确的视觉计数。此外，除了形态学变化之外，许多可以与趋化性迁移一起评估的生理参数通常由于缺乏同时分析方法而未进行评估。我们测试了激光扫描细胞仪（LSC）作为一个方便的平台计数迁移的细胞和并发分析的一些功能与他们的生理status.MethodsWe诱导迁移的THP-1单核细胞与单核细胞趋化蛋白-1或血管内皮生长因子，单独或组合的Nuclepore过滤器的适用性。收集滤膜，将细胞固定在滤膜上，并用核染色剂碘化丙啶染色。趋化性指数通过计算代表性的微观领域，并通过扫描LSC model.ResultsWe发现直接计数和LSC之间的良好相关性的过滤器。此外，LSC仪器中包含的软件工具允许观察迁移细胞中发生的核致密性（碘化丙啶亮度增加）和形态学（核面积和周长增加）的变化。单核细胞趋化蛋白-1和血管内皮细胞生长因子作为添加剂刺激这些参数。ConclusionsLSC分析细胞进行趋化性提供了一个可靠的和全面的评估迁移细胞的数量和分布和他们的一些核参数。该方法可以很容易地扩展到包括一致的分子变化的细胞，由于趋化刺激的评估。© 2005 Wiley利斯公司
BackgroundEvaluation of chemotaxis assays traditionally relies on cumbersome and at times inaccurate visual counting. Moreover, many physiologic parameters that could be evaluated in conjunction with chemotactic migration, aside from morphologic changes, usually are not assessed due to the lack of a simultaneous method of analysis. We tested the suitability of laser scanning cytometry (LSC) as a convenient platform for counting migrated cells and for concurrent analysis of some features associated with their physiologic status.MethodsWe induced migration of THP‐1 monocytes across Nuclepore filters with monocyte chemotactic protein‐1 or vascular endothelial growth factor, alone or in combination. Filters were collected, and cells were fixed on filters and stained with the nuclear stain propidium iodide. Chemotactic indices were obtained by counting representative microscopic fields and by scanning the filters in LSC mode.ResultsWe found an excellent correlation between direct counting and LSC. In addition, the software tools embodied in the LSC instrument allowed the observation of changes in nuclear compactness (increase in propidium iodide brightness) and morphology (increase in nuclear area and perimeter) that occurred in transmigrated cells. Monocyte chemotactic protein‐1 and vascular endothelial growth factor acted as additive stimuli on these parameters.ConclusionsLSC analysis of cells undergoing chemotaxis provides a reliable and comprehensive assessment of the numbers and distribution of migrated cells and some of their nuclear parameters. The method can be easily extended to include the assessment of coincident molecular changes in cells due to chemotactic stimulation. © 2005 Wiley‐Liss, Inc.